Generation of islet-like cells from mouse gall bladder by direct ex vivo reprogramming

Abstract

Cell replacement is an emerging therapy for type 1 diabetes. Pluripotent stem cells have received a lot of attention as a potential source of transplantable β-cells, but their ability to form teratomas poses significant risks. Here, we evaluated the potential of primary mouse gall bladder epithelial cells (GBCs) as targets for ex vivo genetic reprogramming to the β-cell fate. Conditions for robust expansion and genetic transduction of primary GBCs by adenoviral vectors were developed. Using a GFP reporter for insulin, conditions for reprogramming were then optimized. Global expression analysis by RNA-sequencing was used to quantitatively compare reprogrammed GBCs (rGBCs) to true β-cells, revealing both similarities and differences. Adenoviral-mediated expression of NEUROG3, Pdx1, and MafA in GBCs resulted in robust induction of pancreatic endocrine genes, including Ins1, Ins2, Neurod1, Nkx2-2 and Isl1. Furthermore, expression of GBC-specific genes was repressed, including Sox17 and Hes1. Reprogramming was also enhanced by addition of retinoic acid and inhibition of Notch signaling. Importantly, rGBCs were able to engraft long term in vivo and remained insulin-positive for 15 weeks. We conclude that GBCs are a viable source for autologous cell replacement in diabetes, but that complete reprogramming will require further manipulations.

The organoid-initiating cells in mouse pancreas and liver are phenotypically and functionally similar

“…We have recently shown that gall bladder derived epithelial cells can be reprogrammed to become endocrine-like and express insulin using a combination of Pdx1 , Ngn3 and MafA (Hickey et al, 2013). Others have shown that biliary duct cells in the liver can be reprogrammed in vivo and cure diabetes in mice (Akinci et al, 2012).…”

Identification of a candidate stem cell in human gallbladder

“…These cells could also potentially be reprogrammed into hepatocytes and endocrine cells as demonstrated recently (49–51). In addition, since laparoscopic cholecystectomy became clinically available in 1989, the removal of the gallbladder has become routine with 500,000 – 600,000 surgeries every year (52).…”

Novel surface markers directed against adult human gallbladder

“…Aside from the close developmental ontogeny with the pancreas, the functional significance of PDX1 expression in adult human EHBCs is not fully known [9, 11]. It can be speculated that these cells may be more amenable to genetic reprogramming towards the pancreatic endocrine and could be a target for regenerative medicine in diabetes [20, 21]. …”